Herpesviruses are a family of eukaryotic viruses associated with a variety of animal and human diseases, such as orolabial and genital herpes, chicken pox and Epstein-Barr disease. The Herpesvirus family is characterized by its double-stranded DNA and enveloped icosahedral virion, and a genomic size of the double-stranded DNA in the range of 120 to 230 kilobases. The Herpesvirus family is most commonly associated with disease of vertebrates.
Herpesviruses pose a unique challenge to the development of effective vaccines and vaccine administration protocols. These viruses characteristically initially infect epithelial cells on mucosal surfaces. Once a virus has infected an epithelial cell, it then replicates and may spread from cell to cell via intracellular bridges. Herpesviruses are thus capable of establishing a latent infection in neural ganglia and other locations without further exposure to the extracellular environment. Because most herpesvirus infections are initiated on a mucosal surface where systemic immune mechanisms are absent, and spread covertly through intracellular bridges, the generation of systemic immunity by parenteral vaccination has proven ineffective in preventing initial host infection by herpesviruses, although the severity of resulting disease can sometimes be modified through systemic vaccination.
Because of the adverse medical and economic effects from the diseases caused by the herpesviruses, there has been significant effort towards the development of vaccines for these viruses. Originally such efforts focused on whole viral agents, which were inactivated by chemical treatment of the virus or were attenuated by mutation or continuous passage in cell culture to achieve less virulent strains of the virus. Regardless of the outcome of vaccination, these approaches have severe limitations including potential oncogenicity that restrict the use of live or killed herpesviral vaccines in humans. Attenuated herpes virus vaccines cause side effects such as abortion, latency, and disease. Killed virus vaccines are often poorly immunogenic.
The Herpesvirus family, as with many eukaryotic viruses, has characteristic glycoproteins which are carried on the lipid bilayer envelope of the herpesvirus virion. Some of these glycoproteins are believed to function in the initial attachment of virus to cells and penetration of virus into cells. While the glycoproteins themselves do vary to some degree from virus strain to strain, and among the viruses specific to particular hosts, there is a large degree of conservation among the various members of the Herpesvirus family as exemplified by the 45.9% homology between the sequence for bovine herpesvirus 1 (BHV1) glycoprotein I and the human herpes simplex virus type 1 (HSV1) glycoprotein B, as reported by Whitbeck et al., Jour. of Virology, 62:9, pages 3319-3327 (1988). Similar homologies have been found in all other human herpesviruses, herpes simplex type 2, varicella-zoster, Epstein-Barr, cytomegalovirus, and human herpesvirus type 6, as well as all animal herpesviruses yet studied, bovine herpesvirus type 2, pseudorabies, Marek's disease virus, equine herpesvirus type 4, feline herpesvirus, and simian herpesvirus SA8.
Because it has previously been demonstrated that the neutralizing antibody responses produced in vertebrates following challenge with herpesvirus are specific to the glycoproteins carried on the envelope, it has been suggested that vaccines may be developed which include only the glycoproteins, and none of the remaining elements of the viruses necessary for virulence. Accordingly, efforts have been made to characterize, clone, and sequence the glycoproteins for use in creating such vaccines. Such vaccines based on glycoprotein immunogenic agents have also been tested as vaccines both in mammals and in humans. In humans, a vaccine based on a glycoprotein subunit from herpes simplex virus 2 (HSV2) was administered to individuals. Such vaccination elicited antibody titers to the proteins unevenly, but failed to provide effective protection from acquisition of viral infection. Mertz et al., Jour. of Infectious Diseases, 166, pp. 653-660 (1990).
Letchworth and Israel have previously utilized a glycoprotein vaccine from bovine herpesvirus 1, in an attempt to develop an effective vaccine for the BHV1 virus in cattle. Systemic administration of the glycoproteins engendered relatively high titers of serum neutralizing antibodies in the putatively immunized animals. Yet, upon intranasal challenge with virulent virus, these animals showed no resistance to infection or disease. Israel et al., Vaccine 6, pages 349-356 (1988). Thus this and many other herpesviral vaccination studies that were aimed at the generation of systemic neutralizing antibodies consistently proved insufficient to prevent introduction and proliferation of live viruses through the mucosal membranes, the normal portal of entry. In other words, while the vaccinated individual or animal may become less sick, or not sick, because of the vaccination, the individual or animal still becomes a host for the virus and therefore an infectious vector. Obviously such strategy may be useful for particular individuals, but is less than desirable on an epidemiological scale for prevention of wide spread viral disease.
The systemic immune system has been pivotal to the success of many non-herpesviral vaccines. No previous subunit viral vaccines have been focused on the generation or evaluation of the mucosal arm of the immune response. This response is partially independent of the systemic response and is comprised of lymphocytes that circulate between mucosal sites, respond to local antigenic stimuli, and secrete immunoglobulins A and G locally into the mucus. Systemic immunization, via the intravenous, intramuscular, or subcutaneous route does not necessarily evoke a mucosal response. Mucosal immunoglobulins, in contrast to systemic antibodies, are present on the exterior surface of the animal and can neutralize virus before it enters the animal. The mucosal immune system is thus designed to be evoked as the first line of defense against pathogens that enter through the mucosal surface.
In the particular example of a herpesvirus vaccine given in the specification below, the virus in question is Bovine Herpesvirus 1. Bovine Herpesvirus 1 (BHV1) is a ubiquitous pathogenic agent in cattle. As with other herpesviruses, it is an enveloped double-stranded DNA virus. In North America, BHV1 infection causes a respiratory disease known as infectious bovine rhinotracheitis. Infected animals may suffer weight loss, may abort fetuses, may have decreased productivity, and may die from complications of a secondary bacterial pneumonia known as "shipping fever." In Europe, BHV1 has been most commonly manifested as the genital disease infectious pustular vulvovaginitis, but the incidence of respiratory disease is increasing. BHV1 is transmitted by aerosol route, by direct contact, or through semen from infected bulls. While both modified live and killed virus BHV1 vaccines have been used for decades, the disease caused by BHV1 remains a major economic threat to the beef and dairy and artificial insemination industries world wide, since the vaccines have proven ineffective in preventing spread of virulent agents. Since BHV1 establishes latency in the infected hosts, a characteristic typical of herpesvirus in general, recrudescence with subsequent shedding of infectious virus and continued spread of virus to other susceptible animals may occur over the entire lifetime of an animal, even though the animal has an immune response induced by vaccination.
It is quite often the case in modern vaccine design that adjuvants may be introduced with the vaccine to increase immunogenic response to the immunizing agent. This can be the difference between a theoretical and a practical vaccination protocol because soluble glycoproteins are poor immunogens. Traditional adjuvants are formulated to stimulate the systemic and not the mucosal immune response.